Decoding reno-protective mechanisms in mouse Pkhd1 models: Implications for ARPKD therapeutics

解码小鼠 Pkhd1 模型中的肾脏保护机制：对 ARPKD 治疗的影响

• 批准号: 10614404
• 负责人: Ljubica Morizono Caldovic
• 金额: $ 39.27万
• 依托单位: CHILDREN'S RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10614404
• 关键词: ATP12A gene; Address; Affect; Attenuated; Autosomal Dominant Polycystic Kidney; Autosomal Recessive Polycystic Kidney; Biliary; Binding; C-terminal; Cell membrane; Compensation; Cyst; Cystic Kidney Diseases; Data; Disease; Duct (organ) structure; Engineering; Exons; Foundations; Gene Expression; Gene Family; Genes; Genetic; Genetic Models; Genetic Transcription; Human; Infant; Integral Membrane Protein; Kidney; Kidney Diseases; Liver Fibrosis; MYC Family Protein; Messenger RNA; Modeling; Mus; Muscular Dystrophies; Mutant Strains Mice; Mutation; Nuclear; Outcome; Pathogenesis; Pathway interactions; Patients; Perinatal; Phenocopy; Phenotype; Phylogenetic Analysis; Proteins; Regulation; Renal function; Research; Role; Severity of illness; Survivors; Testing; Therapeutic; Transcriptional Regulation; Translating; United States; disease phenotype; experimental study; human disease; in vivo; infancy; insight; integrated circuit; loss of function; mouse genetics; mutant; notch protein; novel; overexpression; promoter; protein protein interaction; renal epithelium; therapeutic target; trafficking

PROJECT SUMMARY There are ~1,500 patients in the United States with autosomal recessive polycystic kidney disease (ARPKD; MIM 173900), a hepatorenal fibrocystic disorder characterized by enlarged kidneys with innumerable collecting duct cysts and progressive loss of renal function. Essentially all cases of ARPKD can be attributed to mutations in PKHD1, which encodes a single-pass transmembrane protein, fibrocystin/polyductin (FPC). We currently have very little insight into the pathogenesis of human ARPKD and thus, treatment is largely supportive. Striking species-specific differences in the PKHD1/Pkhd1 renal phenotype may offer important insights into disease mechanisms. While human patients with either missense or truncating PKHD1 mutations have severe renal cystic disease, mouse Pkhd1 models with engineered truncating mutations (and presumably loss of FPC function) express minimal or no renal disease. Our preliminary studies reveal that while MYC is overexpressed in human ARPKD kidneys, Myc is not overexpressed in mouse Pkhd1 kidneys. In previous studies, we have shown that FPC undergoes Notch-like processing with cleavage of the carboxy terminus (FPC-CTD) from the plasma membrane and nuclear trafficking. Here, we demonstrate that the mouse FPC-CTD binds directly to the Myc P1 promoter and increases Myc expression. Based on these findings, we hypothesize that mouse renal epithelia can compensate for the loss of FPC-CTD function through reno-protective mechanisms and that species-specific, FPC-CTD regulation of Myc expression is central to this reno-protection. We speculate that while these mechanisms are not normally operative in human renal epithelia, they may identify new opportunities for therapeutic targeting in human ARPKD renal disease. We propose two specific aims to test our hypothesis: (1) define how the FPC-CTD regulates Myc/MYC expression in mouse and human renal epithelia and determine whether disruption of the proposed regulatory circuits is a central driver of renal cystogenesis; and (2) compare the FPC-CTD nuclear interactome in mouse and human renal epithelia and test whether differences in transcriptional targets identifies reno-protective pathways in mouse kidneys. Our studies will advance the field by sequentially addressing the transcriptional role of FPC-CTD. Specifically, we will: 1) determine mouse Pkhd1 models; how FPC-CTD related Myc transcriptional regulation contributes to reno-protection in and 2) identify putative mechanisms that allow mouse renal epithelia to compensate for the loss of FPC-CTD nuclear function. Moving forward, these data will lay the foundation for translating mouse reno-protective mechanisms into novel, therapeutic strategies that attenuate human PKHD1-related renal cystic disease.

项目摘要 在美国有大约1,500例常染色体隐性遗传性多囊肾病（ARPKD; MIM 173900），一种肝肾纤维囊性疾病，特征为肾脏增大，伴有无数收集 胆管囊肿和肾功能的进行性丧失。基本上所有的ARPKD病例都可以归因于突变 在PKHD 1中，其编码单次跨膜蛋白纤维囊蛋白/多导管蛋白（FPC）。我们目前有 对人类ARPKD的发病机制了解甚少，因此，治疗主要是支持性的。 PKHD 1/Pkhd 1肾脏表型的显著物种特异性差异可能为以下方面提供重要见解： 疾病机制。尽管具有错义或截短PKHD 1突变的人类患者具有严重的 肾脏囊性疾病，具有工程截断突变的小鼠Pkhd 1模型（以及可能的FPC缺失）， 功能）表达轻微或无肾脏疾病。我们的初步研究表明，虽然MYC在细胞中过度表达， 在人类ARPKD肾脏中，Myc在小鼠Pkhd 1肾脏中并未过表达。在以前的研究中，我们 显示FPC经历Notch-like加工，其中羧基末端（FPC-CTD）从 质膜和核运输。在这里，我们证明了小鼠FPC-CTD直接结合到 Myc P1启动子并增加Myc表达。 基于这些发现，我们假设小鼠肾上皮可以补偿FPC-CTD的损失 通过肾脏保护机制发挥作用，以及Myc表达的物种特异性FPC-CTD调节 是肾脏保护的核心我们推测，虽然这些机制在人类中通常不起作用， 肾上皮细胞，他们可能会发现新的机会，在人类ARPKD肾病的治疗靶向。我们 提出两个具体的目标来验证我们的假设：（1）确定FPC-CTD如何调节Myc/MYC表达 在小鼠和人类肾上皮细胞中，并确定所提出的调节回路的破坏是否是一种 肾脏囊肿形成的中枢驱动因素;以及（2）比较小鼠和人类中的FPC-CTD核相互作用组 肾上皮细胞和测试是否在转录靶点的差异确定小鼠肾脏保护途径 肾脏 我们的研究将通过顺序解决FPC-CTD的转录作用来推进该领域。具体地说， 我们将：1）确定 小鼠Pkhd 1模型; FPC-CTD相关Myc转录调控如何有助于肾脏保护 和2）鉴定允许小鼠肾上皮细胞补偿 FPC-CTD核功能丧失。展望未来，这些数据将为翻译鼠标奠定基础 将肾脏保护机制纳入减轻人PKHD 1相关肾囊肿的新型治疗策略中 疾病

项目成果

Differential regulation of MYC expression by PKHD1/Pkhd1 in human and mouse kidneys: phenotypic implications for recessive polycystic kidney disease.

• DOI: 10.3389/fcell.2023.1270980
• 发表时间: 2023
• 期刊: FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY
• 影响因子: 5.5
• 作者: Harafuji, Naoe;Yang, Chaozhe;Wu, Maoqing;Thiruvengadam, Girija;Gordish-Dressman, Heather;Thompson, R. Griffin;Bell, P. Darwin;Rosenberg, Avi Z.;Dafinger, Claudia;Liebau, Max C.;Bebok, Zsuzsanna;Caldovic, Ljubica;Guay-Woodford, Lisa M.
• 通讯作者: Guay-Woodford, Lisa M.

Transcription factor Ap2b regulates the mouse autosomal recessive polycystic kidney disease genes, Pkhd1 and Cys1.

• DOI: 10.3389/fmolb.2022.946344
• 发表时间: 2022
• 期刊: Frontiers in molecular biosciences
• 影响因子: 5